Long oligodeoxynucleotides: chemical synthesis, isolation via catching-by-polymerization, verification via sequencing, and gene expression demonstration

• Yipeng Yin,
• Reed Arneson,
• Alexander Apostle,
• Adikari M. D. N. Eriyagama,
• Komal Chillar,
• Emma Burke,
• Martina Jahfetson,
• Yinan Yuan and
• Shiyue Fang

Beilstein J. Org. Chem. 2023, 19, 1957–1965, doi:10.3762/bjoc.19.146

• . Structure of phosphoramidites 1a,b and 2, and the catching-by-polymerization (CBP) process. Supporting Information Supporting Information File 2: Experimental details, ODN sequences, PCR primers, and sequence alignments. Acknowledgements Manuscript editing by Dr. S.A. Green, assistance from D.W. Seppala

Cationic oligonucleotide derivatives and conjugates: A favorable approach for enhanced DNA and RNA targeting oligonucleotides

• Mathias B. Danielsen and
• Jesper Wengel

Beilstein J. Org. Chem. 2021, 17, 1828–1848, doi:10.3762/bjoc.17.125

• amine functionalities in their neutral, i.e., non-protonated form. Regarding synthetic strategies, both the use of amine group functionalized phosphoramidites, i.e., functionalized monomeric building blocks, as well as conjugation with amine groups after completion of the ON assembly, are being

• commercially available 3’-phosphoramidite derivative of 5’-O-dimethoxytrityl-2’-O-methyluridine could be converted into an N4-triazole-modified 2’-OMe-cytidine phosphoramidite [60]. This concept was later used to prepare spermine-functionalized 2’-OMe and 2’-O-((2-methoxy)ethyl) (MOE)cytidine phosphoramidites

Double-headed nucleosides: Synthesis and applications

• Vineet Verma,
• Jyotirmoy Maity,
• Vipin K. Maikhuri,
• Ritika Sharma,
• Himal K. Ganguly and
• Ashok K. Prasad

Beilstein J. Org. Chem. 2021, 17, 1392–1439, doi:10.3762/bjoc.17.98

• olefinic nucleoside 106 with mCPBA followed by reaction with 6-chloropurine, N6-(N,N-dimethylformamidine)adenine, or adenine in the presence of K2CO3/NaH in hot DMF afforded the double-headed nucleosides 111–113 (Scheme 26) [30][31][65]. These nucleoside monomers were converted into phosphoramidites and

• phosphoramidites and then introduced into oligonucleotides. The thermal stability of DNA:DNA and DNA:RNA duplexes was determined and it was found that duplexes with a meta-substitution and a phenylacetylene linker were more stable than the corresponding para-substituted and phenyl-linker containing derivatives

• phosphoramidites 160 and 164 which were then incorporated into oligodeoxynucleotides using automated solid phase synthesis. The synthesized oligonucleotides were removed from the solid support by treatment with concentrated aqueous ammonia which resulted in the formation of incorporated monomers 161 and 165 by

Beyond ribose and phosphate: Selected nucleic acid modifications for structure–function investigations and therapeutic applications

• Christopher Liczner,
• Kieran Duke,
• Gabrielle Juneau,
• Martin Egli and
• Christopher J. Wilds

Beilstein J. Org. Chem. 2021, 17, 908–931, doi:10.3762/bjoc.17.76

• chloride, affording both 2'-OMe and 3'-OMe regioisomers. Fortunately, these isomers could be separated by silica gel column chromatography. Other synthetic approaches have since been developed [109][110][111], however, this pioneering work should be appreciated as nowadays, the 2'-OMe phosphoramidites of

DNA with zwitterionic and negatively charged phosphate modifications: Formation of DNA triplexes, duplexes and cell uptake studies

• Yongdong Su,
• Maitsetseg Bayarjargal,
• Tracy K. Hale and
• Vyacheslav V. Filichev

Beilstein J. Org. Chem. 2021, 17, 749–761, doi:10.3762/bjoc.17.65

• -phase DNA or RNA synthesis, the introduction of SaRNA monomers into an RNA backbone involved a 14-step preparation of the phosphoramidite for the SaRNA-TT dinucleotide. Similarly, the incorporation of BCNS groups on a DNA backbone requires the synthesis of thymidine and 2’-OMe-uridine phosphoramidites

Synthesis and properties of oligonucleotides modified with an N-methylguanidine-bridged nucleic acid (GuNA[Me]) bearing adenine, guanine, or 5-methylcytosine nucleobases

• Naohiro Horie,
• Takao Yamaguchi,
• Shinji Kumagai and
• Satoshi Obika

Beilstein J. Org. Chem. 2021, 17, 622–629, doi:10.3762/bjoc.17.54

• technology. In this paper, we describe the synthesis of GuNA[Me] phosphoramidites bearing other typical nucleobases including adenine (A), guanine (G), and 5-methylcytosine (mC). The phosphoramidites were successfully incorporated into oligonucleotides following the method previously developed for the GuNA

• expected to provide further mechanistic insights into how small substituents affect the efficacy and safety of therapeutic oligonucleotides. Thus, the synthesis of GuNA[Me] phosphoramidites bearing other typical nucleobases, i.e., adenine (A), guanine (G), or 5-methylcytosine (mC), instead of the

• immunologically unfavorable cytosine (C), is needed. The preparation of all four phosphoramidites (A, G, mC, and T) is generally not easy because each nucleobase differs in the sensitivity to reactions, and appropriate protecting groups need to be selected [8][21][22][23]. We recently achieved the synthesis of

Changed reactivity of secondary hydroxy groups in C8-modified adenosine – lessons learned from silylation

• Jennifer Frommer and
• Sabine Müller

Beilstein J. Org. Chem. 2020, 16, 2854–2861, doi:10.3762/bjoc.16.234

• , functionalized phosphoramidite building blocks of all four nucleosides are highly desired. The number of commercially available RNA phosphoramidites that carry a suitable functionality for post-synthetic attachment of dyes, reporter groups or other conjugates is still rather limited. In particular, monomer

Nonenzymatic synthesis of anomerically pure, mannosyl-based molecular probes for scramblase identification studies

• Giovanni Picca,
• Markus Probst,
• Simon M. Langenegger,
• Oleg Khorev,
• Peter Bütikofer,
• Anant K. Menon and
• Robert Häner

Beilstein J. Org. Chem. 2020, 16, 1732–1739, doi:10.3762/bjoc.16.145

• individual building blocks (e.g., the phosphoramidites) will enable the creation of a series of molecular probes with different functional groups and varying linker lengths with relatively little effort. The crucial step in the synthetic pathway consists of the conversion of the carbohydrate intermediates

• t-BuOOH, and finally, the protecting groups were removed under basic conditions to give either MPC-1 or MPC-2 as ammonium salts. ETT = 5-(ethylthio)-1H-tetrazole. Preparation of mannosyl phosphoramidites. Starting from 2,3,4,6-tetra-O-acetyl-β-ᴅ-mannopyranose (β-4Ac-Man), the phosphitylation using 2

Electrophilic oligodeoxynucleotide synthesis using dM-Dmoc for amino protection

• Shahien Shahsavari,
• Dhananjani N. A. M. Eriyagama,
• Bhaskar Halami,
• Vagarshak Begoyan,
• Marina Tanasova,
• Jinsen Chen and
• Shiyue Fang

Beilstein J. Org. Chem. 2019, 15, 1116–1128, doi:10.3762/bjoc.15.108

• protections, which contained a tertiary butyl carbamate moiety, were not completely stable under the acidic conditions needed for de-tritylation in each synthetic cycle. Once the protection was lost, in the coupling step, incoming phosphoramidites would react with the free amino groups, and branched ODNs

• comparison with standard ODN synthesis technology, we also synthesized 30a using commercial phosphoramidites and 0.52 µmol 4 (Supporting Information File 2). After purification with RP HPLC, the OD260 of 30a was determined to be 8.30. With these data, we were able to conclude that the dM-Dmoc

• phosphoramidites had similar coupling efficiency as commercial phosphoramidites and the overall yields of ODNs from the dM-Dmoc technology were at the same level of those from standard technologies. The successful synthesis and HPLC purification of the above five ODNs demonstrated that dM-Dmoc is a viable choice

Artificial bioconjugates with naturally occurring linkages: the use of phosphodiester

• Takao Shoji,
• Hiroki Fukutomi,
• Yohei Okada and
• Kazuhiro Chiba

Beilstein J. Org. Chem. 2018, 14, 1946–1955, doi:10.3762/bjoc.14.169

• phosphodiester bond can be an effective linkage not only to construct oligonucleotides but also to conjugate them to various biomolecules, including peptides, sugars, and lipids. The development of a method to activate the supported 5’-terminus, affording useful stable phosphoramidites that are compatible with

Phosphoramidite building blocks with protected nitroxides for the synthesis of spin-labeled DNA and RNA

• Timo Weinrich,
• Eva A. Jaumann,
• Ute M. Scheffer,
• Thomas F. Prisner and
• Michael W. Göbel

Beilstein J. Org. Chem. 2018, 14, 1563–1569, doi:10.3762/bjoc.14.133

• , Max-von-Laue-Str. 7, D-60438 Frankfurt am Main, Germany 10.3762/bjoc.14.133 Abstract TEMPO spin labels protected with 2-nitrobenzyloxymethyl groups were attached to the amino residues of three different nucleosides: deoxycytidine, deoxyadenosine, and adenosine. The corresponding phosphoramidites

• TEMPO-labeled oligonucleotides and of phosphoramidites 5–8. Structures of palindromic oligonucleotides prepared from amidites 5 (22a, 23a), 7 (24a, 25a), and 8 (26a, 27a). Hemiacetals 22b–27b were obtained after photochemical deprotection (custom built apparatus with three LEDs Nichia NCCU033, 365 nm

Oligonucleotide analogues with cationic backbone linkages

• Melissa Meng and
• Christian Ducho

Beilstein J. Org. Chem. 2018, 14, 1293–1308, doi:10.3762/bjoc.14.111

• supra). We have reported that partially zwitterionic NAA-modified DNA oligonucleotides can be obtained by standard solid phase-supported automated DNA synthesis if 'dimeric' phosphoramidite building blocks 49 and 50 are employed (Scheme 5) [72][73]. For the synthesis of 'dimeric' phosphoramidites 49 and

Mechanochemistry of nucleosides, nucleotides and related materials

• Olga Eguaogie,
• Joseph S. Vyle,
• Patrick F. Conlon,
• Manuela A. Gîlea and
• Yipei Liang

Beilstein J. Org. Chem. 2018, 14, 955–970, doi:10.3762/bjoc.14.81

• . Preparation of nucleoside phosphoramidites in a MBM using ionic liquid-stabilised chlorophosphoramidites (route A) or phosphorodiamidites (route B). Preparation of a nucleoside phosphite triester using LAG in a MBM. Internucleoside phosphate coupling linkages in a MBM. Preparation of ADPR analogues using in a

Stimuli-responsive oligonucleotides in prodrug-based approaches for gene silencing

• Françoise Debart,
• Christelle Dupouy and
• Jean-Jacques Vasseur

Beilstein J. Org. Chem. 2018, 14, 436–469, doi:10.3762/bjoc.14.32

• protected by photolabile groups is to use artificial photolabile nucleobases [90]. Generally, these modified nucleobases are introduced into ONs through their corresponding phosphoramidites. Photocaged approaches to inhibit translation: Mikat and Heckel introduced deoxyguanosine and thymidine, respectively

• siRNA–mRNA hybrid, disturbing the cleavage of mRNA by the RISC. Subsequently, Deiters used the same approach with photo 6-nitropiperonyloxymethyl (NPOM)-photocaged siRNAs synthesized from phosphoramidites of the caged uridine and guanosine ribonucleotides (Scheme 20B) [83]. As previously demonstrated

Preparation of trinucleotide phosphoramidites as synthons for the synthesis of gene libraries

• Ruth Suchsland,
• Bettina Appel and
• Sabine Müller

Beilstein J. Org. Chem. 2018, 14, 397–406, doi:10.3762/bjoc.14.28

• advantages of solution chemistry and solid-phase methods. Thus, solid-supported acyl chloride or pyridinium tosylate as the activator of nucleoside-3'-O-H-phosphonates/phosphoramidites, and polystyrene-bound trimethylammonium periodate as oxidation reagent have been demonstrated to be superior for dimer and

• to 49% [23]. 5. Phosphitylation and coupling of trinucleotide synthons in solid phase DNA synthesis To be used as building blocks in standard phosphoramidite synthesis, fully protected trimers need to be converted in phosphoramidites (Figure 7). This has been described in a number of reports [19][22

• diisopropylamine, which is neutralized by benzylmercaptotetrazole released back after the reaction. The tetrazole derivative is sufficiently acidic to act as a scavenger for diisopropylamine converting it into the ammonium salt. Thus, fully protected trimers can be converted to phosphoramidites without the loss of

Aminosugar-based immunomodulator lipid A: synthetic approaches

• Alla Zamyatina

Beilstein J. Org. Chem. 2018, 14, 25–53, doi:10.3762/bjoc.14.3

Synthesis of oligonucleotides on a soluble support

• Harri Lönnberg

Beilstein J. Org. Chem. 2017, 13, 1368–1387, doi:10.3762/bjoc.13.134

• the nucleobases are usually protected with acyl groups and the 5´-OH of the monomeric building block with a 4,4’-dimethoxytrityl group (DMTr), or sometimes with its monomethoxytrityl analog (MMTr) [4][5]. To achieve coupling, phosphoramidites are activated with azoles [6], such as tetrazole [7], its

• the 3´-terminal hydroxy function unprotected and, hence, available for one step conversion to a phosphoramidite building block [45]. Such phosphoramidites are widely used for the assembly of ODNs useful in protein engineering by oligonucleotide directed mutagenesis [46][47][48][49]. Cleavage of the

• extraction with water. The couplings were carried out with 1.5 equiv of standard 2-cyanoethyl-N,N-diisopropylphosphoramidites in THF or MeCN, using DCI as an activator. Unreacted phosphoramidites were quenched by EtOH and the support was precipitated before the oxidation step, repeatedly when needed. The

Facile synthesis of a 3-deazaadenosine phosphoramidite for RNA solid-phase synthesis

• Elisabeth Mairhofer,
• Elisabeth Fuchs and
• Ronald Micura

Beilstein J. Org. Chem. 2016, 12, 2556–2562, doi:10.3762/bjoc.12.250

• mechanisms. Therefore, 1-deazaadenosine (c1A), 1-deaza-2’-deoxyadenosine (c1dA), 3-deazaadenosine (c3A), and 3-deaza-2’-deoxyadenosine (c3dA), and the corresponding phosphoramidites to prepare oligoribonucleotides are highly requested nucleoside modifications. Unfortunately, synthetic approaches to achieve

• them are troublesome and time consuming, in particular for c3A. To the best of our knowledge, only two papers have reported the synthesis of c3A phosphoramidites so far [16][17]. Thereby, the major bottleneck is access to the naked nucleoside. Although the c3A nucleoside is commercially available

DNA functionalization by dynamic chemistry

• Zeynep Kanlidere,
• Oleg Jochim,
• Marta Cal and
• Ulf Diederichsen

Beilstein J. Org. Chem. 2016, 12, 2136–2144, doi:10.3762/bjoc.12.203

• Phosphoramidites 4 and 11 were introduced at position seven of 13mer oligonucleotides ON1 and ON2 applying automated solid-phase synthesis (Table 1 and Supporting Information File 1, Table S1). The last step in the oligonucleotide synthesis involved the deprotection of the amine group using ammonium hydroxide at

Terminal lipophilization of a unique DNA dodecamer by various nucleolipid headgroups: Their incorporation into artificial lipid bilayers and hydrodynamic properties

• Emma Werz and
• Helmut Rosemeyer

Beilstein J. Org. Chem. 2015, 11, 913–929, doi:10.3762/bjoc.11.103

• oligonucleotides (LONs 10–15), each terminally lipophilized with different nucleolipid head groups, were synthesized using the recently prepared phosphoramidites 4b–9b. The insertion of the LONs within an artificial lipid bilayer, composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and 1-palmitoyl

• lipophilization and studies on the interactions of lipophilized nucleic acids with lipid bilayers is of growing importance. As lipophilic head groups we used a series of nucleolipid cyanoethyl phosphoramidites that were recently synthesized [13][14][15][16][17]. Results and Discussion Figure 1 indicates the

• -, sesqui-, and diterpenes as well as single and double-chained alkyl groups (completed by an alicyclic alkyl group), have been introduced for the lipophilization of the pyrimidine nucleosides. The resulting nucleolipids were converted into their corresponding 2-cyanoethyl phosphoramidites 4b–9b. Figure 3

An unusually stable chlorophosphite: What makes BIFOP–Cl so robust against hydrolysis?

• Roberto Blanco Trillo,
• Jörg M. Neudörfl and
• Bernd Goldfuss

Beilstein J. Org. Chem. 2015, 11, 313–322, doi:10.3762/bjoc.11.36

• , in Pd catalysts. Keywords: chirality; hydrolysis; phosphorus; rearrangements; terpenoids; Introduction Phosphorus halides are highly reactive intermediates for the synthesis of phosphites and phosphoramidites [1][2][3][4][5], which are widely used, for example, as ligands in catalysts [6][7][8][9

TEMPO-derived spin labels linked to the nucleobases adenine and cytosine for probing local structural perturbations in DNA by EPR spectroscopy

• Dnyaneshwar B. Gophane and
• Snorri Th. Sigurdsson

Beilstein J. Org. Chem. 2015, 11, 219–227, doi:10.3762/bjoc.11.24

• increased flexibility of UC and UA, inspection of the line-shape of their CW EPR spectra reveals subtle differences between the four base-pairing partners A, T, G and C when placed in a DNA duplex. Results and Discussion Synthesis of TA, UA, UC and their corresponding phosphoramidites The spin-labeled

• UC-containing oligonucleotides The phosphoramidites of TA (5), UA (10) and UC (14) were used to incorporate the spin-labeled nucleosides into DNA oligonucleotides using solid-phase synthesis [77]. The low stability of the nitroxide functional group in the TEMPO moiety towards acids lead to almost ca

NAA-modified DNA oligonucleotides with zwitterionic backbones: stereoselective synthesis of A–T phosphoramidite building blocks

• Boris Schmidtgall,
• Claudia Höbartner and
• Christian Ducho

Beilstein J. Org. Chem. 2015, 11, 50–60, doi:10.3762/bjoc.11.8

• blocks 6 (Figure 1) [38]. Overall, 24 different oligonucleotide sequences with one to four NAA-modifications at various positions were synthesized. The stereochemistry of the NAA-motif was either (S) or (R) (obtained by application of the corresponding phosphoramidites (S)-6 or (R)-6 for DNA synthesis

• NAA-modified DNA oligonucleotides [38], thus making the NAA-linkage an interesting structural motif for oligonucleotide analogues. Using 'dimeric' T–T phosphoramidites 6, it was only possible to introduce the NAA-modification at T–T motifs. While this was fully sufficient for initial studies, it will

• be of major importance to develop methods for the synthesis of NAA-modified oligonucleotides with the NAA-motif at more variable positions in the base sequence. The synthesis of 'dimeric' X–T phosphoramidites (X = A, C, G) would enable an introduction of the NAA-linkage at every position in an

Nucleic acid chemistry

• Hans-Achim Wagenknecht

Beilstein J. Org. Chem. 2014, 10, 2928–2929, doi:10.3762/bjoc.10.311

• phosphordiesters or -triesters. Finally, this approach using phosphoramidites as nucleoside building blocks was significantly further developed in 1981 by Beaucage and Caruthers [4]. Since then, oligonucleotides of up to 50-mers in length have become available by an extremely efficient solid-phase methodology that

Autonomous assembly of synthetic oligonucleotides built from an expanded DNA alphabet. Total synthesis of a gene encoding kanamycin resistance

• Kristen K. Merritt,
• Kevin M. Bradley,
• Daniel Hutter,
• Mariko F. Matsuura,
• Diane J. Rowold and
• Steven A. Benner

Beilstein J. Org. Chem. 2014, 10, 2348–2360, doi:10.3762/bjoc.10.245

• synthesis from six phosphoramidites (four standard and two AEGIS). They were then mixed in equal amounts, heated and cooled. The 3’-fragments were then extended at 48 °C using Phusion DNA polymerase to give a nicked construct and the nicks were sealed with ligase. The first indication that the GACTSB AEGIS